Polyamine is a general term for those alkylamines with two or more amino groups. There are four types of polyamines [putrescine (H2N(CH2)4NH2), cadaverine (H2N(CH2)5NH2), spermidine (H2N(CH2)4NH(CH2)3NH2) and spermine (H2N(CH2)3NH(CH2)4NH(CH2)3NH2)] and acetylated forms thereof in the human body.
Relatively recently, it was found that two types of diacetylpolyamines [N1,N8-diacetylspermidine (hereinafter expressed as “DiAcSpd”) and N1,N12-diacetylspermine (hereinafter expressed as “DiAcSpm”)] are excreted in urine though very small in quantities. While these components occupy only 1.4% and 0.6% of the total polyamine, respectively, in the urine of healthy persons, the ratios of these components remarkably increase in the urine of cancer patients as compared to other polyamine components. Further, it has been shown that these components have other characteristics of tumor markers (Sugimoto, M. et al., J. Cancer Res. Clin. Oncol., 121, 317-319 (1995); Hiramatsu, K. et al., J. Cancer Res. Clin. Oncol., 123, 539-545 (1997)).
Initially, DiAcSpd and DiAcSpm were quantitatively determined by a method which was a combination of a fractionation system by HPLC and a detection system using enzyme (Hiramatsu, K. et al., J. Biochem., 117, 107-112 (1995)). However, more simple measuring methods have been developed. In particular, with respect to the measurement of DiAcSpm, an ELISA method using a specific antibody was developed recently (Hiramatsu, K. et al., J. Biochem., 124, 231-236 (1998)). However, preparation of a kit containing an ELISA measuring system has not been achieved yet.
Techniques for measuring DiAcSpm by immunoassay have also been developed (Japanese Patent Unexamined Publication No. H11-75839; Japanese Patent Unexamined Publication No. 2000-74917), but they have room for improvement in terms of measurement sensitivity and cross-reactivity.
It is well known that metabolism of polyamines are activated in association with cell proliferation. In fact, polyamine contents tend to increase in various cancer tissues as compared to normal tissues. Since any of these polyamines is contained abundantly in actively proliferating tissues and is excreted in large quantity in the urine of cancer patients as compared to the urine of healthy persons, they are evaluated as tumor markers. A kit which is capable of measuring the total polyamine content easily by an enzymatic determination method without discriminating acetylated polyamines and free polyamines in urine and without discriminating the four types of polyamines in urine has already been commercialized (Labo-Search PolyamineAuto; AIT Corporation). This kit is used as one of clinical test methods. However, with respect to urinary total polyamine, it has become clear that a relatively large number of false negative results are found in malignant tumor patients and that total polyamine significantly increases in association with not only malignant tumors but also with various diseases and conditions such as inflammatory diseases, cardiac infarction, cirrhosis, process of curing of wounds, etc. Thus, it is considered that total polyamine cannot be evaluated as a marker specific to malignant tumors (Shunichiro Kubota, NIPPON-RINSHO (Japan Clinical Medicine) 53, Special Issue, pp. 501-505 (1995)). On the other hand, researchers are now revealing that some of the problems total polyamine has can be avoided by measuring diacetylpolyamines (DiAcSpd and DiAcSpm) discretely (Sugimoto, M. et al., J. Cancer Res. Clin. Oncol., 121, 317-319 (1995)).